Carnitine has an important function in beta-oxidation of fatty acids in every organ of a living body. Cells in every organ produce ATP (adenosine triphosphate), which is the energy source for the cells, in mitochondria owing to the beta-oxidation of fatty acids. An acyl group of the fatty acids is transported in the mitochondria through a carnitine action to be transformed into acyl Co-A, and then the acyl Co-A is beta-oxidized to generate ATP. Many carnitines are diet-induced, and, though a part thereof is biosynthesized in the liver and the brain, almost all the cells need to uptake carnitines from outside; therefore, it has been assumed that a carnitine-specific transporter exists in the cell membrane according to past physiological studies.
Studies have been conducted on the carnitine uptake by various cells by the use of experiment systems employing the removed organ perfusion method or the isolated cell membrane vesicular system. However, it has been difficult to analyze the system of carnitine transport through cell membrane in detail by the conventional methods, and there has been a demand for isolation and analysis of the transporter itself.
The rat CT1 (Sekine, T. et al., Biochem. Biophis. Res. Commun., Vol. 251, pp. 586-591, 1998) has been isolated as the carnitine transporter. The hOCTN2 (Tamai, I. et al., J. Biol. Chem., Vol. 273, pp. 20378-20382, 1998) has been reported to be a human homologous gene of the CT1. The carnitine transporters are widely distributed in organs of the whole body to contribute to the carnitine uptake by various cells. They exist also in the kidney to carry out a function of reabsorption of carnitine from the renal tubules.
According to the past studies, it has been clarified that a carnitine concentration in the testis and the epididymis is the highest in the living body and reached the 2,000 times of the serum level depending on the specie of the carnitine. Since the duct of epididymis is an organ governing nutrition and maturity of sperms, the relationship between carnitine and sperm function has attracted attention.
It has been clarified that the sperms produced in the testis are premature and low in motility and lacks in fertilizability at that stage. The sperms leaving the testis move to the epididymis and subjected to various modifications therein and matured. Further, flagellar movement is necessary for the sperm fertility, which requires enormous energy. It is reasonable to consider that a large amount of carnitine is required for the production of ATP which is the energy source for the flagellar movement.
Actually, a study revealing a positive relationship between the carnitine concentration in the human semen and the number and motility of sperms has been reported. Also, in the clinical medicine, it has been reported that carnitine administration had resulted in an increase in fertilization probability of idiopathic infertility patients.
Thus, the carnitine transport specific to the testis and the epididymis has been considered to have an important function in the maturity and the fertilizability of sperms and has been studied by the experiment systems employing the removed organ perfusion method or the isolated cell membrane vesicular system. Though the existence of an active carnitine transport system has been clarified from the studies, its molecular substance has been indefinite.
Organic anion transporters OAT1 (organic anion transporter 1) (Sekine, T. et al., J. Biol. Chem., Vol. 272, pp. 18526-18529, 1997), OAT2 (Sekine, T., FEBS letter, Vol. 429, pp. 179-182, 1998), OAT3 (Kusuhara, H. et al., J. Biol. Chem., Vol. 274, pp. 13675-13680, 1999), and OAT4 (Cha, S. H. et al., J. Biol. Chem., Vol. 275, pp. 4507-4512, 2000), which have the central role in drug transport in the kidney, liver, brain, and placenta, have been isolated and reported by some of the inventors of the present invention. These transporters belonging to an OAT family are transporters that can transport many different organic anions in chemical structure and also transport of various anionic drugs.
Recently, an OCT (organic cation transporter) family which recognizes organic cations also has recently been identified. Since carnitine is an organic ion having both of a positive charge and a negative charge, it is assumed that the carnitine transporter is positioned between the OAT family and the OCT family from the point of molecular evolution.
In view of these facts, the present inventors have presumed that the carnitine transporter specific to the testis and the epididymis belongs to the organic ion transporter family.